MAY
1961
NOTES
1G77
A similar transformation employing 16a-methyl- medium and a 55-hr. incubation time with steroid were prednisone as the substrate gave a less polar prod- used, 1.95 g. of 16a-methylprednisone was transformed. Chromatography of the concentrated chloroform extract 224 mcc ( E 8,650). The structure of over silicic acid afforded by chloroform elution 0.114 g. of uct (11),";:A: I1 was assigned as lGa-methyl-l-pregnene-l7a,21- tan powder which ran as a single compound, slightly faster diol-3,11,20-trione because the observed ultra- moving than 16a-methylprednisone, in the Shull system,lO The powder was rechromatographed over Florisil and eluted violet maximum was in good agreement with with 1%methanol and 2% methanol in methylene chloride. the predicted value6 and by analogy with the The combined crystalline fractions on recrystallization from proved structure of I. acetone-hexane gave 0.083 g. of 11, m.p. 206-212", [a]? 224mp ( e 8650), 'A$': 2 . 8 8 ' ~(OH), Qualitative evidence for the same type of +116" (dioxane),"":::A: transformation was also obtained with 1,4-preg- 5.88 p (11- and 20-carbonyl), 6.00 p (3-carbonyl), 6.22 p nadiene lla117a,21 - triol - 3,20 - dione which (AI). Anal. Calcd. for CzZH3006: C, 70.56; H, 8.08. Found: was transformed into a less polar product with an C, 70.83; H, 8.03. ultraviolet maximum a t 237 m p (AA = - 11 rnp'). INSTITUTE OF MICROBIOLOGY Similar qualitative results were also obtained with RUTGERS UNIVERSITY 16a-methyl-1,4-pregnadiene-l1 a,17a,21-triol-3,20NEW BRUNSWICK, N. J. dionee0
-
SCHERINQ CORP. BLOOMFIELD, N. J.
EXPERIMENTAL'
l-Pregnene-l7~,dl-diol-5,11,dO-lrione (I). T o each of four 250-ml. Erlenmeyer flasks waa added 50 mi. of 1% yeast extract-dextrose medium and an inoculum of spores of Estrogens. 111. Synthesis of Streptomyces sp. W 3808 (Waksman collection). After a 644-Methylequilenin'i hr. incubation with rotary shaking a t 30" the cultures were each transferred to one of four 2-1. Erlenmeyers containing 400 ml. of the same medium. Following another 48 hr. of TADL. PATTON incubation, 0.130 g. of prednisone was added to each flask. After 112 hr. of additional incubation the contents of all Received July 88,1960 flasks were pooled, the mycelium was removed by filtration, and a water wash of the mycelium waa added to the filtrate. Procedures for the introduction of a methyl The combined filtrate was extracted with chloroform and the extracts were concentrated to a small volume. The group a t positions-l,2, 4, and 6 of estrone and estraresulting solution was chromatographed over silicic acid and diol and a t position-l of equilenin have been rethe column was eluted with chloroform. - ~is the purpose of this note to report The combined crystalline fractions ( I ) from the chro- p o r t e d . ' ~ ~It 225 mp a procedure fbr the introduction of a methyl group matogram (0.070 g . ) melted a t 185-189", "A:J: ( e 6600) and gave a positive test with red tetrazolium a t position-4 of equilenin. reagent. In the Shull system10 I migrated slightly faster Since the phenolic rings of estrone and estradiol than cortisone. The infrared spectrum of I was essentially condense with formaldehyde and secondary amines the same as that of l-pregnene-17~,21-diol-3,11,20-trione,4 lv5 the pheto yield 2-dialkylaminomethylestrogens, but I was clearly impure. $1-Acetate of I. A solution of 0.050 g. of I in 1.0 ml. of nolic ring of equilenin would be expected to react pyridine and 1.0 ml. of acetic anhydride was allowed to similarly. In fact, it was found that formaldehyde stand a t room temperature overnight. Excess water was and morpholine condense with equilenin to give added and the resulting precipitate (0.048 9.) was collected by filtration. Recrystallization from aqueous methanol and one compound, a monosubstituted product, which from acetonc-hexane gave 0.023 g. of the 21-acetate of I, was assigned the structure of 4-morpholinomethylm.p. 235-239", CY]^ $131" (acetone), 225 mp equilenin (I). This assignment was made because ( e 8400). Mattox and Kendall5 report I-pregnene-lia,21equilenin is a 5,6-disubstituted 2-naphthol, and diol-3,11,20-trione 21-acetate, m.p. 245-246", [ a ] +138" 225 mp ( L 9100). The infrared spectrum the reaction would be expected to occur a t the (acetone),"?:A: of the 21-acetate of I matched that of an authentic sample. position equivalent to the one a t which it occurs 16a-Alelhyl-1-pregnene- 17a,B1-diol-3,11,20-lrzone (11). in 2-naphthol. The latter has been shown to undergo Under essentially the conditions employed in the preparation of I, with the exception that a 2% yeast extractdextrose
a.
(6) Calculated for the shift in U.V. AA = X (l-pregnene17a,21-dioI-3,11,20-trione 21-acetate)-A (prednisone) = 225
-
A (16a-methylmp - 238 mp = -13 mp; observed A (11) prednisone) = 225 mp - 238 mp = -13 mp. ( 7 ) The preparation of 1,4-pregnadiene-lla,l7a,21-triol3,20-dionc is given in U. S. Patent 2,837,464. The ultraviolet maximum (methanol) is a t 248 mp (e 17,800). (8) E. P. Oliveto et al., J . A m . Chem. SOC., 80, 4431 (1958). (9) All melting points are corrected. Analyses and optical data were obtained by the Physical Chemistry Departments of these laboratories. The infrared spectra were interpreted by Mr. Richard Wayne. (10) G. M. Shull, Abstracts of the 126th Meeting of the American Chemical Society, New York, 1954, p. DA.
(1) The preceding paper in this series is T. I,. Patton, J . Org. Chem., 25,2148 (19GO).
( 2 ) This investigation was supported by a grant, CY2873, from the National Cancer Institute, U. S. Public Health Service. (3) C. Djerassi, G. Rosenkranz, J. Romo, J. Pataki, and S. Kaufmann, J . A m . Chem. Soc., 7 2 , 4540 (1950). (4) J. Iriarte and H. J. Ringold, Tetrahedron, 3, 28 (1958). (5) T. L. Patton, Chem. & Ind. (London), 923 (1959). (6) H. Dannenberg, C. H. Deering, and D. Donnenbergvon Dresler, Hoppe-Seyler's 2. physiol. Chem., 317, 174 (1959). (7) E. Velarde, J. Iriarte, H. J. Ringold, and C. Djerassi, J . Org.Chem., 24,311 (1959).
1678
NOTES
VOL.
2G
TABLE I ULTRAVIOLET ABSORPTION MAXIUAOF SUBSTITLTED ~-~\~AIWTHOI,S Compound
n ~ (pf n ~ a u ) ~ '
2-Taphthol 1 -1Iorpholinomethyl%naphthol Efiuilenin
I I1 a
336(91,194) 230 (61,044)
265(3911) 269(3890)
275(4550) 279(4890)
28G(X%0l) 290 (3850)
320( 1861) 324(2150)
,331 (216:3)
231 (65,476) 236(57,320) 237 (67,330)
270(4428) 274(4510) 274 (4400)
281 (5047) 284(5510) 285 (5480)
292(3571) 296(4510) 297 (4360)
328(1752) 333 (2600) 333 (25GO)
311 (641) 345(3120) 345 (2880)
335 (2180)
The solvent was 95% ethyl alcohol.
the hlannich reaction specifically at position-1 ;8,9 therefore, by comparison, equilenin should react specifically a t position-4, the counterpart of position-1 in 2-naphthol. I n addition, the introduction of the morpholinomethyl group into equilenin causes a hathochromic shift of the ultraviolet spectrum which is almost identical to the spectral shift observed when the same group is introduced into position 1 of 2-naphthol (see first four lines of Table I). The hydrogenolysis of I with Raney nickel by a known procedure1f5 removed the morpholine part of the molecule to give 4methylequilenin (11). The ultraviolet absorption spectrum of I1 is essentialli parallel to that of I (see Table I ) ; therefore, the aromatic structure remained unchanged throughout the hydrogenolysis reaction. Evidence that the carbonyl group at C-17 was not reduced during the hydrogenolysis reaction was provided by the infrared spectrum of I1 which showed a strong absorption peak at 5 . 7 7 ~In . addithis is chartion, I1 exhibited absorption a t 2.99~; acteristic of the phenolic hydroxyl group. The infrared absorption spectrum of I, like the spectra of 2-dialkylnminomethylestrogens,1did not show absorption in the 3 p region. EXPERIMENTAL'^ 4-Morpholinon~ethylequilenin (I). Equilenin (266 mg. 0.001 mole) was dissolved in a solution of ethyl alcohol ( I 2 ml.) and morpholinc ( 0 . 3 ml.) hy heating; then 37% formaldehyde (0.1 nil.) was added. After the so!ution had refluxed for 30 min. an additional 0.2 ml. of formaldehyde was added, and the solution was heated at reflux temperatiire for 2 hr. It then remained at room temperature overnight. The solvent was removed at water pump pressure,
(8) J. Decombe, Compt. rend., 197, 258 (1933). ( 9 ) F. F. Blicke, Org. Reactions, 1, 303 (1942). (10) IIelting points were taken on a FisherJohn8 block a:id are Lincorrectrd. The microanalyses were done by Dr.
Carl Tie 'cke, Ln1)oratory of Microchemistry, Teaneck, S . J. A Beckman LIodel 1)U spectrophotometer with a photomiiltinlier attachment mas used to obtain the ultraviolet snrctra. The infrared s,wctra were recorded by a Perkin-Elmer Model 2 1 infrared spectrophotometer using a so !ium chloride prism.
and the viscous residue was made to solidify by the addition of a few drops of ethyl alcohol. The crude product was taken up in ether and washed with 10% hydrochloric acid to remove the amino steroid. The acid wash was cooled and made alkaline with ammonium hydroxide. The product was extracted with several portions of ether. After washing the combined extracts with water and drying them over anhydrous sodium sulfate the ether was evaporated a t reduced pressure. The residue was recrystallized from ethyl alcohol to give 235 mg. of pink colored crystals. The second recrystallization gave colorless crystals of analytic1 purity, m.p. 167-168"; 5.75 p (carbonyl a t (31,) and 7.14 p (free methylene at CI6). Anal. Calcd. for C23H2703N. C, 75.58; H, 7.44; IT, 3.83. Found. C, 75.61; H, 7.39; N, 3.91. 4-Methylequilenin (11). A suspension of Rsney nickel ( 1 9.) in a solution of I (121 mg., 0.00033 mole) in 9570 ethyl alcohol (11 ml.) was heated at reflux temperature for 15 hr. During this time (overnight) the suspension apparently humped rather vigorously, because a large amount of the material had heen lost through the top of the condenser and was on the outside of the condenser, flask, and heating mantle. The product which remained inside the apparatus was isolated as follows. The suspension was diluted with ethyl alcohol and filtered hot to remove t h e catalyst. The filtrate was concentrated to a volume of less than 1 ml. and set aside to allow crystallizntion to occur. The product weighed 20 mg. ( 7 . 1 7 , ) : m.p. 244-246' (red melt). Normally, the vield of I1 would be expected to be 60-65% since this yield of 2-methylestradiol is obtained by the hydrogenolysip of 2-diethylaminomethylestradiol under the same reaction conditions.' However, this expectation remains unconfirmed, because the rarity of the starting material prevented the repetition of this experiment. Further recrystallization from ethyl alcohol gave 1 1 mg. of analytically pure TI, m.p. 749-251' (red melt); 2.99 p (OH a t C3), 5.77 p (carbonyl a t C,,), and 7.15 p (free methylene a t C1d. Anal. Calcd. for C19H2&: C, 81.40; H, 7.19. Found: C, 81.62; H , 7.30. 1-Mor~holinomethvl-I-naphthol. This was synthesized by a known procedure t o give a product melting a t 115' (lit l 1 m.p. 115- 116').
Acknowledgment. The author wishes t o acknomledge a gift of equilenin generously supplied by Dr. W. L. Glen of Ayerst, McKenna & Harrison, Ltd., Montreal, Canada. SECTION OF EXPERIMENTAL MEDICINE THE UNIVERSITYOF TEXASM. D. ANDERSON HOSPITAL A N D TUMOR INSTITUTE 25, TEX. HOUSTON (11) R. L. Shriner, G. F. Grillot, and 1%'. 0. Teeters,
J. Am. Chem. SOC.,68, 946 (1946).
